Graduation date: 1997

The gut hormone glucagon-like peptide-2 (GLP-2) has numerous beneficial effects on the intestinal epithelium, including increased mucosal growth and proliferation. GLP-2 is also necessary for the adaptive intestinal re-growth that occurs upon re-feeding after fasting. Although insulin-like growth factor (IGF)-1 and the IGF-1 receptor are known to be required for GLP-2-induced crypt-cell proliferation, the precise cellular localization of the IGF-1 receptor through which the intestinotrophic actions of GLP-2 are mediated remains unknown. I hypothesized that small intestinal growth responses to GLP-2 occur through an intestinal epithelial IGF-1 receptor-dependent pathway, through the use of an inducible, intestinal epithelial-specific IGF-1 receptor knockout (IE-igf1rKO) mouse. Intestinal growth and proliferative responses were examined in IE-igf1rKO and control mice following treatment with GLP-2, as well as in animals that were fasted and re-fed to induce GLP-2-dependent adaptation. In Chapter 3, it was demonstrated that IE-igf1rKO mice, as compared to control littermates, had normal small intestinal weight, morphometric parameters, proliferative index and differentiated epithelial cell lineage distribution. Administration of GLP-2 for 30 minutes increased nuclear translocation of !-catenin in non-Paneth crypt-cells, and stimulated the crypt-cell proliferative marker c-Myc 90 minutes following GLP-2 treatment, in control littermates but not in IE-igf1rKO mice. In Chapter 4, adaptive re-growth was studied by fasting IE-igf1rKO and control animals for 24 hours, or by fasting and then re-feeding mice for 24 hours. Small intestinal weight, crypt depth, villus height and crypt-cell proliferation were decreased in both control and IE-igf1rKO mice after 24 hour fasting. While re-feeding in control mice restored all of these parameters, re-fed IE-igf1rKO mice displayed abrogated adaptive re-growth of the crypt-villus axis as well as reduced crypt-cell proliferation. In Chapter 5, control mice responded to chronic GLP-2 with increased small intestinal weight, mucosal cross-sectional area, crypt depth, villus height and crypt-cell proliferation. However, the GLP-2-induced increase in crypt-cell proliferation was absent in IE-igf1rKO mice, in association with impaired growth of the crypt-villus axis. Taken together, these results indicate that the proliferative responses of the intestinal epithelium to exogenous GLP-2 administration and during conditions of GLP-2-dependent adaptive re-growth are dependent on the intestinal epithelial IGF-1 receptor.

The gut hormone glucagon-like peptide-2 (GLP-2) has numerous beneficial effects on the intestinal epithelium, including increased mucosal growth and proliferation. GLP-2 is also necessary for the adaptive intestinal re-growth that occurs upon re-feeding after fasting. Although insulin-like growth factor (IGF)-1 and the IGF-1 receptor are known to be required for GLP-2-induced crypt-cell proliferation, the precise cellular localization of the IGF-1 receptor through which the intestinotrophic actions of GLP-2 are mediated remains unknown. I hypothesized that small intestinal growth responses to GLP-2 occur through an intestinal epithelial IGF-1 receptor-dependent pathway, through the use of an inducible, intestinal epithelial-specific IGF-1 receptor knockout (IE-igf1rKO) mouse. Intestinal growth and proliferative responses were examined in IE-igf1rKO and control mice following treatment with GLP-2, as well as in animals that were fasted and re-fed to induce GLP-2-dependent adaptation. In Chapter 3, it was demonstrated that IE-igf1rKO mice, as compared to control littermates, had normal small intestinal weight, morphometric parameters, proliferative index and differentiated epithelial cell lineage distribution. Administration of GLP-2 for 30 minutes increased nuclear translocation of !-catenin in non-Paneth crypt-cells, and stimulated the crypt-cell proliferative marker c-Myc 90 minutes following GLP-2 treatment, in control littermates but not in IE-igf1rKO mice. In Chapter 4, adaptive re-growth was studied by fasting IE-igf1rKO and control animals for 24 hours, or by fasting and then re-feeding mice for 24 hours. Small intestinal weight, crypt depth, villus height and crypt-cell proliferation were decreased in both control and IE-igf1rKO mice after 24 hour fasting. While re-feeding in control mice restored all of these parameters, re-fed IE-igf1rKO mice displayed abrogated adaptive re-growth of the crypt-villus axis as well as reduced crypt-cell proliferation. In Chapter 5, control mice responded to chronic GLP-2 with increased small intestinal weight, mucosal cross-sectional area, crypt depth, villus height and crypt-cell proliferation. However, the GLP-2-induced increase in crypt-cell proliferation was absent in IE-igf1rKO mice, in association with impaired growth of the crypt-villus axis. Taken together, these results indicate that the proliferative responses of the intestinal epithelium to exogenous GLP-2 administration and during conditions of GLP-2-dependent adaptive re-growth are dependent on the intestinal epithelial IGF-1 receptor.

The purpose of this study was to look at the genotoxic and cytotoxic effects of benzo-á-pyrene (BáP), a chemical mutagen that is present in cigarette smoke, on the insulin-like growth factor-I (IGF-I) gene. Women who smoke during pregnancy are more likely to have a growth-restricted baby. We hypothesized that BáP exerts its effects through genotoxic and cytotoxic avenues. The cytotoxicity is manifested by chromosomal abnormalities and a decrease in the rate of cell division. The genotoxicity is manifested by changes in certain genes known to be important in mammalian fetal development such as IGF-I. IGF-I is implicated in intrauterine growth restriction (IUGR), a problem that greatly increases the risk of perinatal morbidity and mortality. To futher understand the mechanism by which BáP influences the normal growth and development of human placental cells, human placental trophoblast cells from an established immortalized cell line were utilized. Cells were cultured in appropriate media, starved (using starvation "Serum Free Medium"), and treated with two doses of BáP, 1µM (dose 1) and 5µM (dose 2). Chromosomes were prepared for cytogenetic analysis and visualized using light microscopy after Giemsa staining. Chromosomal aberrations were identified and the rate of cell division was determined through the analysis of the mitotic index for treated cells compared to a control group. To further understand the influence of BáP on the IGF-I gene expression level, RNA was extracted from control and treated cells, from which cDNA was synthesized and used for further analysis using polymerized chain reaction (PCR). The PCR results were used to better understand the genotoxicity of BáP, while chromosomal aberration analysis was used to determine the cytotoxic effects of BáP on human placental cells. Our results indicate that many chromosomal abnormalities were present in the treated groups compared to the control group. In addition, there was a significant decrease in the mitotic index of the BáP-treated cells (MI=0.3%) verses the control group (MI=0.93%), p value 0.0447. Through the PCR assay, we speculate that there is a dose-related response to BáP of the IGF-I RNA expression level, with low levels in the treated groups compared to the control group. We conclude from these results that BáP influences placental cells at both the gene and chromosome level. It also affects the cell cycle of human placental cells. It is known that smoking is deleterious for fetal development. We believe that the current study brings us closer to understanding the mechanism by which smoking can lead to fetal growth restriction.

Thesis (Ph. D.)--Auburn University, 2008. / Abstract. Vita. Includes bibliographical references (p. 62-77).

Thesis (Ph. D.)--Cornell University, January, 2007. / Vita. Includes bibliographical references.

Thesis (M.S.)--Rutgers University, 2009. / "Graduate Program in Microbiology and Molecular Genetics." Includes bibliographical references (p. 51-61).

Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 117-126).

published_or_final_version / Medicine / Master / Master of Research in Medicine

published_or_final_version / abstract / toc / Obstetrics and Gynaecology / Doctoral / Doctor of Philosophy